Sophorae Subprostratae Radix Extract for Prevention and Treatment of Respiratory Diseases

ABSTRACT

The present invention relates to an extract isolated from Sophorae Radix useful for prevention and treatment of respiratory diseases. More specifically, the present invention relates to a pharmaceutical drug comprising the Sophorae Radix extract having excellent effects of inhibiting airway contraction, respiratory infections, 5-lipoxygenase activity, phosphodiesterase 4 activity, airway hyper-responsiveness and airway remodeling; antagonistic activity against leukotriene D 4 ; and an antitussive effect, thus being useful for prevention and treatment of respiratory diseases such as asthma, acute or chronic bronchitis, allergic rhinitis, acute upper respiratory infections and acute lower respiratory infections, and the like.

FIELD OF THE INVENTION

This invention relates to Sophorae Radix extract useful for preventionand treatment of respiratory diseases. This invention also relates to apharmaceutical drug comprising Sophorae Radix extract useful forprevention and treatment of respiratory diseases due to their excellenteffects in preventing airway contraction, respiratory infection,5-lipoxygenase, phosphodiesterase 4, airway hyper-responsiveness orairway remodeling, leukotriene D4, as well as having an antitussiveeffect.

BACKGROUND OF THE INVENTION

Asthma is a typical respiratory disease having repetitive andspasmodically symptoms of difficulty in breathing, coughs and stridor.About 30% pediatric asthma patients start to show the symptoms withinone year from birth while about 80% of asthma patients show the symptomsat the age of 4-5. In Korea, the rate of asthma outbreak is about 10%.

The incidence rate of asthma varied to some extent depending on thecountry, race, age and the like. According to the British report in1991, about 7% of adults and 13.5% of children are suffering fromasthma. In Korea, the number of asthma patients is on the increase dueto the drastic change in life styles as well as in environmentalconditions such as severe pollution and accumulated stress. The age ofdeveloping asthma has been lowered recently due to environmentalcontamination such as pollution and its symptoms have been prolonged.

Respiratory obstruction as one of characteristics of asthma occurs via 3steps: i.e., contraction of bronchial smooth muscle, tylosis ofpulmonary mucosa, and accumulation of sticky mucus in bronchi andbronchioles. Of them, contraction of bronchial smooth muscle is rathereasily recovered.

In the attack of extrinsic(allergic) asthma, it is known that IgE playsa very important role and IgG is also often involved. IgE releasesmediators (histamine, SRS-A, ECF-A, NCF, PAF, Kinin, PGs, etc.) whichinduce a hypersensitivity reaction by activating mast cells. The causeof intrinsic(non-allergic) asthma is still not known but this kind ofasthma appears to be mediated by autonomic nerves. In an intrinsicasthma patient, a cholinergic stimulus can directly release mediatorssuch as histamine from mast cells, increase secretion of goblet cells,dilate pulmonary blood vessels, and contract trachea, bronchi, and largebronchioles, thereby causing bronchial spasm and increasing release ofmucus.

So far there is no cure for asthma. Although there are many methods anddrugs which have been used for the prevention of spasm and complicationsdue to asthma they have not been satisfactory. One of the most effectiveways of preventing the attack of asthma may be to find the very factorsthat are involved in causing asthma. Examples of therapeutic agents thathave been used to treat asthma are inhaling bronchodilator drugs, oralor injectable bronchodilator drugs (sympathetic stimulators andtheophyllines), steroid preparations (inhaling, oral and injectableform, etc.), leukotriene antagonists (montelukast, pranlukast, zileuton,etc.), anti-allergic drugs (cromolyn disodium, ketotifen, etc.) and thelike.

Bronchitis can be either acute or chronic. According to its causes,bronchitis is divided into allergic, infectious, and extrinsicbronchitis, while pathologically it is classified catarrhalis,suppurative, occlusive, ulcerative, and infiltrative bronchitis. Themost frequent cause of bronchitis is due to infection with bacteria,viruses, fungi and the like. People who normally not infected with theabove pathogens can be infected when their systemic immune system getsweakened. Allergic bronchitis can be a direct allergic reaction due toinhalation of allergens or a partial symptom due to a generalizedallergic reaction. Extrinsic bronchitis may occur due to a chemicalstimulus such as chlorine and sulfur dioxide gas, or due to a physicalstimulus such as dusts. People living in large cities with polluted aircan be readily exposed to respiratory infections. In case of acutebronchitis, from the pathological point of view, it is easy to observeruber, swelling, and xerosis and also mucous or suppurative secretions.In general, asthmas can be recovered without incurring complications.However, if it is progressed into a chronic asthma, it results inswelling, tylosis and atrophy. In a prolonged chronic asthma, it resultsin fiber proliferation, bronchostenosis or pulmonary emphysema. The mostpeculiar symptoms of bronchitis are coughs and phlegm. If the cause ofbronchitis is due to infection, there often develops fever and chestpain, whereas if it is due to extrinsic factors, there often developsirritations on the mucus of mouth, nose, eye, etc. In therapy, cough isconsidered as a sort of a bodily defense and thus it is not recommendedto intentionally stop coughing. It is essential that a therapy for thecause be conducted along with other measures. In winter, it is desirableto increase room temperature and administer a small amount of codeine,atropine, ephedrine, antihistamine agents, and the like. Use of steroidsor theophyllines for treating infections is not satisfactory.

Nasal allergic inflammation often refers to nasal allergy or allergicrhinitis. It entails symptoms of sudden continuous coughs, release of alarge amount of clear nasal mucus, stuffy nose, heavy head, release oftears, and the like. When the symptoms are similar but the allergens arenot identified it is coryza vasomotoria. For example, the above symptomsmay occur when body temperature is temporarily lowered in the morningand they are usually recovered within a few hours, and these symptomsare commonly seen in people in a cold season. This can be sometimesconfused with nasal cold but it differs from the cold and oftenaccompanies asthma and hives.

The allergic reaction in allergic rhinitis is an antigen-antibodyhypersensitivity reaction, wherein histamine is released from mast cellsand cell walls of basophils, and arachidonic acid is released to produceprostaglandins and leukotrienes by cyclooxygenase (COX) and5-lipooxygenase(5-LO), thereby mediating the initial reaction occurringbetween 2-90 min after being exposed to an antigen and the post reactionoccurring 4-8 hrs thereafter. The initial reaction is proceeded with bya mediating substance while the post reaction is mediated by cellinfiltration. Further, allergic and non-allergic rhinitis both serve asrisk factors for developing asthma.

In therapies, desensitization is performed when the antigen is clearlyidentified. Other therapies such as use of drugs, surgeries, physicaltherapies but they are not considered as a complete cure.

There are various respiratory infections due to such as pyogenicbacteria, special bacteria (Mycobacterium tuberculosis, Corynebacteriumdiphtheriae, spirochete, etc.) viruses, fungi, and they are also dividedinto acute and chronic respiratory infections. When nasal cavity,pharynx, and larynx are independently infected the diseases are calledby their respective organ names. However, when the above organs areinfected as a whole they are called as upper airway infections, and arepresenting example is upper airway disease. Besides, in the event ofinfections due to special bacteria or fungi, they are also frequentlycalled as upper airway tuberculosis, upper airway diphtheriae, upperairway candida and the like.

As stated above, respiratory diseases such as asthma, allergic rhinitis,acute and chronic bronchitis differ with respect to their causes andsymptoms but they have common characteristics in the following fewaspects.

First, they are all inflammatory diseases. These respiratory diseasesare caused by allergies, infections, etc., but inflammation plays acrucial role in exacerbation and treatment of the diseases. That is,introduction of leukocytes stimulated by allergies, infections, etc.,into a respiratory tract and activation therein and the variouscytokines released from leukocytes and inflammatory mediatorsdeteriorate diseases and affect the therapeutic treatment.

Second, contraction and relaxation of the respiratory tract does notperform normally thus making respiration difficult. That is, therespiratory tract is impaired thereby performing an excess reaction(asthma) in response to a normal stimulus or it becomes too narrowed toperform a normal bronchial respiration thus requiring an appropriatetreatment.

Third, in major drug therapies, antiinflammatory agents, agents thatinhibit respiratory contraction, bronchodilators, agents that inhibitrespiratory release play important roles and other therapeutic drugs arealso commonly used in combination. For example, anti-histamine drugs,anti-cholinergic drugs, beta 2 receptor agonist, steroids, leukotrieneD4 receptor antagonists, phosphodiesterase 4 inhibitor of theophyllinesare commonly used. Nevertheless, bronchodilator drugs such asanti-cholinergic drugs, beta 2 receptor agonist, etc., are not effectivein treating inflammation but they simply alleviate the symptoms.Therefore, long-term use of the drugs may cause drug resistance andthere is also a risk of exacerbation. Steroids which are known effectivein treating inflammation but they have serious side effects and are notsuitable for long term use and also shown not effective in treatingchronic bronchitis. Therefore, the above two drugs have been prescribedto be combined for administration but the steroids drug has beenformulated in an inhalation form rather than as one for oraladministration due to its adverse effects thus lowering its compliancedue to the difficulty in administration. Therefore, there is a need todevelop a novel therapeutic drug which can resolve the above-mentionedlimitations in the currently used therapeutic drugs and effectivelyimprove the symptoms. However, as stated above, various leukocytes andvarious cytokines and inflammatory mediators are involved in respiratorydiseases, it is difficult to treat the respiratory diseases with asingle ingredient chemical and thus a natural extract having variousactive ingredients and mechanism may be able to serve as an effectivetherapeutic drug.

Further, there appear to be many causes for respiratory diseases such asasthma, bronchitis, allergic rhinitis, acute lower respiratoryinfections(bronchitis, bronchiolitis, etc.), acute upper respiratoryinfection(tonsillitis, pharyngolaryngitis) but they are treated only fortemporary release, and there is usually a problem of recurrence of thediseases after treatments. Therefore, prevention and treatment ofrespiratory diseases has been raised as one of the most important tasksto fulfill in medical science and the development of a novel therapeuticdrug for the fundamental prevention and treatment of respiratorydiseases is in urgent need.

Sophorae Radix of the present invention to be used as a crude drug is ashrub with a height of 1-2 m. It has about 2-5 cylindrical roots withyellowish brown color. Its stem has a cylindrical shape, has a groove onthe surface and is densely covered with short and soft hairs where theupper part of the stem is normally bent in the form of

a Chinese letter. Its roots, which are used as drugs after drying, havea long cylindrical shape and are a bit bent with a length of 10-25-35cm, a diameter of 0.3-1 cm. Its surface is brown or dark brown, hasvertically-formed wrinkles and lengthy lenticels with a bit of rotation.Its main place of product is Gwangseo Province in China. In orientalmedicines, it has been used to treat tumors, edema, pains, jaundice,diarrhea, hemorrhoid, and the like. Its active ingredients are alkaloidssuch as matrine, oxymatrine, anagyrine, methylcytisine and the like andflavonoids such as sophoranone, sophoradin, sophoranochromene,sophoradochromene and the like (Chinese Medicine Encyclopedia, JungdamPublishing, pp. 2627-2632, 1998). However, the effect of Sophorae Radixextract on respiratory diseases has not been studied yet.

SUMMARY OF THE INVENTION

The inventors of the present invention have conducted extensive effortsto develop a therapeutic drug effective in treating respiratorydiseases, and as a result, have discovered that Sophorae Radix extracthas excellent activities of inhibiting airway contraction, respiratoryinfections, 5-lipoxygenase, phosphodiesterase 4, airwayhyper-responsiveness and airway remodeling; antagonistic activityagainst leukotriene D4; and antitussive effect. Therefore, an object ofthe present invention is to provide a therapeutic drug for preventionand treatment of respiratory diseases comprising Sophorae Radix extractas an active ingredient.

BRIEF DESCRIPTION OF THE DRAWINGS

The aforementioned aspects and other features of the present inventionwill be explained in the following description, taken in conjunctionwith the accompanying drawings, wherein:

FIG. 1 is a graph showing the effect on the airway hyper-responsiveness[SAL: saline, OVA: ovalbumin, SOS: extract prepared in PreparationExample 2];

FIG. 2 A-F are pictures showing the effect of the extract prepared inPreparation Example 2 (SOS) or montelukast on airway mucus expression inlung tissues of OVA-sensitized and -challenged mice;

FIG. 3 is a histogram showing the effect of the extract prepared inPreparation Example 2 (SOS) or montelukast on airway mucus expression inlung tissues of OVA-sensitized and -challenged mice;

FIG. 4 A-F are pictures showing the effect of the extract prepared inPreparation Example 2 (SOS) or montelukast on α-smooth muscle expressionin lung tissues of OVA-sensitized and -challenged mice;

FIG. 5 is a histogram showing the effect of the extract prepared inPreparation Example 2 (SOS) or montelukast on α-smooth muscle expressionin lung tissues of OVA-sensitized and -challenged mice;

FIG. 6 A-F are pictures showing the effect of the extract prepared inPreparation Example 2 (SOS) or montelukast on peribronchial fibrosis inlung tissues of OVA-sensitized and -challenged mice;

FIG. 7 is a histogram showing the effect of the extract prepared inPreparation Example 2 (SOS) or montelukast on fibrosis in lung tissuesof OVA-sensitized and -challenged mice;

FIG. 8 is a histogram showing the effect of the extract prepared inPreparation Example 2 (SOS) or montelukast on total lung collagencontent of OVA-sensitized and -challenged mice; and

FIG. 9 is a result of Western blot showing the effect of the extractprepared in Preparation Example 2 (SOS) or montelukast on IL-4, IL-5,and IL-13 protein expression in lung tissues of OVA-sensitized and-challenged mice.

DETAILED DESCRIPTION OF THE INVENTION

In one aspect, the present invention relates to Sophorae Radix extractuseful for prevention and treatment of respiratory diseases. In anotheraspect, the present invention relates to a pharmaceutical drugcomprising Sophorae Radix extract as an active ingredient which haveexcellent effects of inhibiting airway contraction, respiratoryinfections, 5-lipoxygenase activity, phosphodiesterase 4 activity,airway hyper-responsiveness, and inhibitory activity of airwayremodeling; antagonistic activity against leukotriene D4; antitussiveeffect; therefore being useful for prevention and treatment ofrespiratory diseases such as asthma, acute and chronic bronchitis,allergic rhinitis, acute lower respiratory infection(bronchitis,bronchiolitis, etc.), acute upper respiratory infection(sphagitis,tonsillitis, laryngitis) and the like.

The present invention is described in further detail hereunder.

The method of preparing Sophorae Radix extract according to the presentinvention is as follows.

1) Crude Sophorae Radix is extracted via reflux by using about 7 to 10times of water or an alcohol solution with reference to the weight ofSophorae Radix and then filtered. The remnant is extracted again byadding about 4 to 7 times of water or an alcohol solution with referenceto the weight of the combined Sophorae Radix, followed by heating, andthen filtered. Thus obtained two filtrates are combined together andfiltered.

2) The filtrate obtained in the above step 1) undergoes phase separationby using an equal amount of water-saturated low grade alcohol or anonpolar solvent and then concentrated under reduced pressure at 50-60°C.

3) The above concentrate undergoes azeotropic concentration by addingabout 20 to 50 times of water with reference to the total weight of theconcentrate obtained in the above step 2), is uniformly suspended withan equal amount of water and is then placed under lyophilization.

More specifically, the original crude Sophorae Radix is added with wateror an alcohol solution and extracted under reflux for 2 to 5 hrs,wherein the amount of water or an alcohol solution is preferably about 7to 10 times with reference to the weight of crude Sophorae Radix. Then,the above extract is filtered. The filtrate is then added with about 4to 7 times of water or an alcohol solution with reference to the weightof the crude Sophorae Radix, heated, reextracted for 2-5 hrs, filteredand combined with the previously obtained filtrate thereby increasingthe extraction efficiency.

Here, if the amount of water is too little it cannot provide efficientstirring and also lower the solubility of the resulting extract thusdecreasing the extraction efficiency, whereas if the amount of water isin excess it will increase the amount of low grade alcohol and thenonpolar solvent to be used in the following purification step thusbeing uneconomical and also raises a handling problem.

In the present invention, the extraction consists of the firstextraction and the second extraction. When a crude extract is producedin bulk it usually contains a large amount of water content due to thecrude drug itself although filtration is performed efficiently.Therefore, the present invention prevents the relatively low extractionefficiency obtained by the first extraction only. In addition, a furtherstudy on the extraction efficiency by the inventors of the presentinvention showed about 80-90% of the total extract is obtained by thesecond extraction, which suggests that additional extractions of morethan second extraction is not necessary and also uneconomical.

The extract obtained by the first and the second extractions as statedabove is filtered, concentrated and then purified to get rid ofunnecessary impurities such as proteins, polysaccharides, fatty acidsand the like. In the present invention, the filtrate is added with anequal amount of low grade alcohol or a nonpolar solvent to perform phaseseparation 2-4 times thereby obtaining solvent fraction while separatingimpurities. Here, examples of low grade alcohols include alcohol havingcarbon atoms of from 1 to 6, preferably butyl alcohol, propyl alcohol,or isopropyl alcohol. Examples of nonpolar solvents are ethylacetate,dichloromethane, chloroform, carbon tetrachloride or methylethylketone.When the amount of the low grade alcohol or the nonpolar solvent is lessthan that of the filtrate it often produces granules due to the presenceof impurities such as fatty acids and thus the phase separation cannotbe performed effectively and also the extraction efficiency on theactive ingredients becomes relatively low thus being not efficient. Thefraction of a low grade alcohol or a nonpolar solvent obtained as aresult of the above phase separation is concentrated under reducedpressure at 50-60° C. to remove the solvent remaining in the specimen.Thus obtained concentrate undergoes azeotropic concentration 2-3 timeswith about 25-50 times of water with reference to the total weight ofthe concentrate, and then uniformly suspended by adding an equal amountof water. The main reason of performing azeotropic concentration is toeffectively control the content of the remaining low grade alcohol inorder to use the extract of the crude drug as a raw material for apharmaceutical drug.

Further, the extract of Sophorae Radix of the present invention can beobtained, in addition to the above-mentioned method, by extracting withwater, water-saturated low grade alcohol or a nonpolar solvent followedby purification. Examples of low grade alcohols include alcohol havingcarbon atoms of from 1 to 6, preferably butyl alcohol, propyl alcohol,or isopropyl alcohol. Examples of nonpolar solvents are ethylacetate,dichloromethane, chloroform, carbon tetrachloride or methylethylketone.

Thus obtained Sophorae Radix extract undergoes lyophilization and thenfinal extract is obtained in powder form. This final extract hasexcellent activities of inhibiting airway contraction, respiratoryinflammation, 5-lipoxygenase activity, phosphodiesterase 4 activity,airway hyper-responsiveness and airway remodeling; antagonistic activityagainst leukotriene D4, antitussive effect, etc., and is thus expectedto be useful for the prevention and treatment of respiratory diseases.

The Sophorae Radix extract of the present invention can be administeredin various oral and parenteral forms during clinical studies. When theyare formulated diluents such as a filler, a bulking agent, a binder, awetting agent, a disintegrating agent, a surfactant and the like or anexcipient are used.

Solid preparations for oral administration include tablets, pills,powders, granules, capsules, troches, suppositories and the like. Thesesolid preparations are prepared by adding at least one excipientselected from the group consisting of starch, calcium carbonate, sucroseor lactose, gelatin, and the like to a mixture of lignan and lactonecompound or its derivative. Besides, in addition to a simple excipient,a lubricant such as magnesium stearate, talc, and the like can be used.Bases for suppositories are hard fat triglyceride esters, polyethyleneglycol, polysorbate, cacao oil, laurin butter, glycerol, gelatin and thelike.

Liquid preparations for oral administration are suspensions, solutions(syrups, drinks, etc.), emulsion and the like. For example, variousexcipients such as wetting agents, sweeteners, flavoring agents, andpreservatives can be used in addition to the most frequently useddiluents such as water and liquid paraffin.

Preparations for parenteral administration are sterilized solutions,non-aqueous solutions, suspensions, emulsions, lyophilizers. Solventsfor non-aqueous solutions, suspensions or emulsions are vegetable oils,propylene glycol, polyethylene glycol, olive oil, and ethyl oleate.

Sophorae Radix extract of the present invention has been used in folkremedies for long time and its safety has been confirmed by toxicitytest. The dosage of the Sophorae Radix extract depends on variousfactors such as the rate of body absorption, body weight, age, sex,health conditions, diets of a subject and time required foradministration, method of administration, excretion rate, seriousness ofdiseases, and the like. As shown in pharmacological experiments, it ispreferable to administer Sophorae Radix extract about 1-15 mg/kg of bodyweight. Therefore, the Sophorae Radix extract of the present inventionto be used as an active ingredient of a pharmaceutical drug should bemanufactured considering the effective range of pharmaceuticalefficacies, and thus manufactured pharmaceutical preparations in theform of unit formulation can be administered at regular intervals and/oraccording to a specialized medication committed under the supervision ofa medical specialist or by the request of a subject.

The present invention will be described in more detail with reference tothe following examples, however, they should not be construed aslimiting the scope of the present invention.

PREPARATION EXAMPLE 1 Preparation of Sophorae Radix Extract

250 g of Sophorae Radix minced to a size of about 1.0 cm were well mixedand added with 2 L of water and then heat-extracted for 5 hrs whilestirring. The resulting filtrate was collected and the remnant was addedwith 1.5 L of water and heat-extracted for 3 hrs. The above twofiltrates were combined and then concentrated to the final volume of 1.5L. The above concentrate was added with an equal volume ofwater-saturated n-butyl alcohol and performed phase separation 3 times.Only n-butyl alcohol fraction was collected and concentrated underreduced pressure at 58° C. until the extract becomes dry. After most ofthe n-butyl alcohol and water are evaporated, it was added with 0.1 L ofwater and performed azeotropic concentration 3 times. The resultant wasresuspended in an equal volume of distilled water and then lysophilizedto finally obtain Sophorae Radix extract in powder.

PREPARATION EXAMPLE 2 Preparation of Sophorae Radix Extract

Sophorae Radix was cleaned with water and dried. 250 g of the SophoraeRadix was added with 2 L of 50%(v/v) ethanol solution and extractedunder reflux for 6 hrs while stirring. The resulting filtrate wascollected and the remnant was added with 1.5 L of 30%(v/v) ethanolsolution and heat-extracted for 3 hrs. The above two filtrates werecombined and then concentrated to the final volume of 1.5 L. The aboveconcentrate was added with an equal volume of water-saturated n-butylalcohol and performed phase separation 3 times. Only n-butyl alcoholfraction was collected and concentrated under reduced pressure at 58° C.until the extract becomes dry. After most of the n-butyl alcohol andwater are evaporated, it was added with 0.2 L of water and performedazeotropic concentration 3 times. The resultant was resuspended in anequal volume of distilled water and then lysophilized to finally obtainSophorae Radix extract in powder.

PREPARATION EXAMPLE 3 Preparation of Sophorae Radix Extract

Sophorae Radix was cleaned with water and dried. 250 g of the SophoraeRadix was added with 2 L of water-saturated butyl alcohol solution andextracted under reflux for 6 hrs while stirring. The resulting filtratewas collected and the remnant was added with 1.5 L of water-saturatedbutyl alcohol solution and heat-extracted for 3 hrs. The above twofiltrates were combined and then concentrated under reduced pressure at58° C. until the extract becomes dry. After most of the n-butyl alcoholand water are evaporated, it was added with 0.2 L of water and performedazeotropic concentration 3 times. The resultant was resuspended in anequal volume of distilled water and then lysophilized to finally obtainSophorae Radix extract in powder.

EXAMPLE 1 Experiment on Inhibition of Airway Contraction (In Vitro)

To evaluate the inhibitory effect of Sophorae Radix extract prepared inPreparation Examples 1-3 against airway contraction, experiments wereperformed using ablated bronchi as described below and the results areshown in Table 1.

[Method]

A Hartely male guinea pig(400-450 g, SLC, Japan) was sensitized byintravenously injecting 1.5 mL/kg of anti-ovalbumin anti-serum. 48 hrsafter the sensitization, the guinea pig was killed by exsanguinationsand then its trachea were isolated. Other tissues attached to thebronchi were removed in Krebs-Heseleit solution and the bronchi were cutout in the form of a ring so that it contains 2-3 cartilages. Whilemaintaining bronchial muscles intact, the cartilage parts of the ringwere cut out and connected with thread on both sides and hung in anorgan bath. After stabilization, it was added to induce a maximumcontraction by adding 10 μg/mL of carbachol. The bronchi were washedwith Krebs-Heseleit solution and stabilized. Indomethacin (2 μmoles) wasadded and the test material ‘X’ was added into organ bath one minutelater. In 5 min, 10 μg/mL ovalbumin(OVA) was added to inducecontraction. Rate of airway contraction was calculated by comparingcontractions induced by carbachol and OVA. The airwaycontraction/relaxation was measured by using a physiological activitymeasuring device (MP150, BioPAC system) connected to a force transducer(FT4, BioPAC system).

TABLE 1 Inhibition Rate of Classification Concentration BronchialContraction (%) Preparation Example 1 0.25 mg/mL 42.4 0.5 mg/mL 76.8Preparation Example 2 0.25 mg/mL 48.8 0.5 mg/mL 83.0 Preparation Example3 0.25 mg/mL 43.8 0.5 mg/mL 80.6 Positive Montelukast 10 μM 24 Control

As shown in the above Table 1, it was confirmed that the Sophorae Radixextract of the present invention has an effect of inhibiting airwaycontraction.

EXAMPLE 2 Experiment on Inhibition of Airway Contraction (In Vivo)

To evaluate the inhibitory effect of Sophorae Radix extract prepared inPreparation Examples 1-3 against airway contraction, experiments wereperformed as described below by exposing antigens to a sensitized guineapig and the results are shown in Table 2.

[Method]

A Hartely male guinea pig(400-450 g, SLC, Japan) was sensitized byintravenously injecting 1.5 mL/kg of anti-ovalbumin anti-serum. 48 hrsafter the sensitization, the guinea pig was administered with a drugorally. In 30 min, the guinea pig was pretreated by injecting 10 mg/kgof pyrilamine maleate, 10 mg/kg of indomethacin, and 0.1 mg/kg ofpropranolol subcutaneously, respectively. Then, the guinea pig wasplaced in Double Chamber Plethysmograph Box(HSE, Germany) installed withplethysmometer for measuring various respiratory indices and measuredbasic airway resistance values. 30 min after the pretreatment, 1% OVAwas nebulized for 2 min in the form of aerosol by preparing a compressedair under high pressure. Airway resistance was measured for 30 min as anindicator of bronchospasm.

TABLE 2 Inhibition Rate of Classification Amount (mg/kg) Bronchospasm(%) Preparation Example 1 100 30 200 52 400 75 Preparation Example 2 10040 200 55 400 70 Preparation Example 3 100 35 200 53 400 71 PositiveMontelukast 10 45 Control 40 75

As shown in the above Table 2, it was confirmed that the Sophorae Radixextract of the present invention has an effect of inhibiting airwaycontraction in a sensitized guinea pig.

EXAMPLE 3 Experiment on Inhibition of Airway Infection

To evaluate the inhibitory effect of Sophorae Radix extract prepared inPreparation Examples 1-3 against bronchial inflammation, experimentswere performed as described below by utilizing the increase inleukocytes such as eosinophils to the area of pulmonary bronchi byexposing antigens to a sensitized mouse and the results are shown inTable 3.

[Method]

A 6 week old BALB/c female mouse (SLC, Japan) was sensitized byadministering 0.2 mL of a mixture consisting of 10 μg of OVA and alumintraperitoneally at Days 0, 7 and 14, respectively. 8 and 10 days afterthe last sensitization, 0.7% OVA was sprayed to the mouse for 50 min inthe form of aerosol by a compressed air under high pressure to induceairway inflammation. 24 hrs after the induction of the airwayinflammation, the bronchialveolar were washed with 1.5 mL of a phosphatebuffer solution. The washed solution was collected and the number ofleukocytes and eosinophils in the washed solution were counted,respectively. Further, white blood cell infiltration into tissues,tissue impairments, etc., was observed by means of hematoxylin and eosinstaining in the lung tissues and they were scaled accordingly. SophoraeRadix extract was orally administered 7-10 days after the lastsensitization.

TABLE 3 Inhibition Rate of Classification Amount (mg/kg) AirwayInfection (%) Preparation Example 1 200 25 400 55 Preparation Example 2200 30 400 50 Preparation Example 3 200 28 400 54 Positive Montelukast10 24 Control 40 25

As shown in the above Table 3, it was confirmed that the Sophorae Radixextract of the present invention has an effect of inhibiting airwayinfection. While the effect of montelukast as a positive controlexhibits saturation of pharmaceutical efficacy at a relatively low levelwithout dosage-dependent effect, the Sophorae Radix extract of thepresent invention is shown to have sufficient pharmaceutical effectwhich is also dosage-dependent.

EXAMPLE 4 Inhibition of 5-lipoxygenase (5-lipoxygenase, 5-LO) Activity

To evaluate the inhibitory effect of Sophorae Radix extract prepared inPreparation Examples 1-3 against a leukotriene-producing enzyme, a majorcause of asthma, experiments were performed as described below and theresults are shown in Table 4.

[Method]

Human peripheral blood mononulear leukocyte (PBML) was stabilized inHank's balanced salt solution (HBSS) at 37° C. and then added withSophorae Radix extract and allowed to react for 15 min. To the abovemixture, arachidonic acid was added as a base material and producedleukotriene B4 (LTB4) for 15 min. The amount of thus produced LTB4 wasmeasured by using Enzyme immuno assay (EIS) kit.

TABLE 4 Concentration Inhibition Rate of Classification (mg/mL)5-Lipoxygenase Activity (%) Preparation Example 1 0.03 97 0.1 100Preparation Example 2 0.03 98 0.1 100 Preparation Example 3 0.03 98 0.1100 * positive control NDGA(Nordihydro-guaiaretic acid) IC₅₀ = 87.4 nM

As shown in the above Table 4, it was confirmed that the Sophorae Radixextract of the present invention has an effect of inhibiting5-lipoxygenase activity.

EXAMPLE 5 Inhibition of Phosphodiesterase 4 (PDE4) Activity

To evaluate the inhibitory effect of Sophorae Radix extract prepared inPreparation Examples 1-3 against a phosphoesterase 4, a major cause ofasthma, experiments were performed as described below and the resultsare shown in Table 5.

[Method]

Human U937 cells were stabilized in the mixed solution comprising 50 mMTris-HCl and 5 mM MgCl₂ (pH 7.5) at 25° C. Sophorae Radix extract and1.01 μM of ([3H]cAMP+cAMP) as a base material were combined together andallowed to react for 20 min to produce adenosine. The amount of thusproduced adenosine was quantitated by measuring the amount of[3H]adenosine.

TABLE 5 Concentration Inhibition Rate of Classification (mg/mL)phosphoesterase 4 (%) Preparation 0.03 78 Example 1 0.1 100 Preparation0.03 76 Example 2 0.1 100 Preparation 0.03 79 Example 3 0.1 100 *positive control IBMX(3-isobutyl-1-methylxanthine) IC₅₀ = 20.7 μM

As shown in the above Table 5, it was confirmed that the Sophorae Radixextract of the present invention has an effect of inhibitingphosphoesterase 4 activity.

EXAMPLE 6 Antagonistic Activity against Leukotriene D4 Receptor (LTD4)

To evaluate the inhibitory effect of Sophorae Radix extract prepared inPreparation Examples 1-3 against a LTD4 receptor, a major cause ofasthma, experiments were performed as described below and the resultsare shown in Table 5.

[Method]

LTD4 receptor isolated from the pulmonary tissues of a Duncan Hartelyguinea pig was stabilized in 50 mM Tris-HCl buffer solution (5 mM CaCl₂,5 mM MgCl₂, 100 μg/mL bacitracin, 1 mM benzamidine, 0.1 mMphenylmethylsulfonyl fluoride) at 25° C. Then, Sophorae Radix extractand 0.2 nM [3H]LTD4 were added to the above mixture and allowed toreact. Binding rate was analyzed via Radioligand binding assay, andnon-specific binding was determined by means of 0.1 μM LTD4. Specificbinding rate 85%, Kd 0.2 nM, Bmax 0.24 pmol/mg protein.

TABLE 6 Concentration Inhibition Rate of Classification (mg/mL) LTD4Receptor (%) Preparation 0.03 78 Example 1 0.1 89 Preparation 0.03 81Example 2 0.1 89 Preparation 0.03 82 Example 3 0.1 88 * positive controlLTD4 IC₅₀ = 1.12 nM

As shown in the above Table 6, it was confirmed that the Sophorae Radixextract of the present invention has an effect of inhibiting LTD4receptor activity.

EXAMPLE 7 Antitussive Effect

The method used was the method by M. H. Boskabady et al. (journal ofEntnopharmacology 97, 2005, 79-82) with a bit of modifications.

A Hartley male guinea pig (450˜500 g, SLC, Japan) was orallyadministered with a drug. 1 hr after the administration, the guinea pigwas placed in Double Chamber Plethysmograph Box (HSE, Germany) installedwith plethysmometer and stabilized by allowing a 5 min of time foradaptation. After the stabilization, 0.6M citric acid was sprayed to theguinea pig for 7 min in the form of aerosol by preparing a compressedair under high pressure. The guinea pig was observed continuously by atrained observer and the number of coughs induced by the above spray ofcitric acid aerosol was measured by using a microphone and a speaker.The coughs were distinguished from a normal sneeze due to its particularsigns such as a peculiar and high sound with its mouth open, movement ofabdomen, and an instant change in air flow.

TABLE 7 Dose Drug (mg/kg) Cough Inhibition (%) CMC 1 12.3 ± 2.6 (N = 7)Preparation Example 1 400  7.8 ± 3.7 (N = 5) 37 Preparation Example 2200 10.6 ± 2.6 (N = 8) 14 400  6.6 ± 1.4 (N = 9) 47 400 (2 h)  8.7 ± 2.3(N = 6) 29 Preparation Example 3 400  8.4 ± 1.5 (N = 5) 32 Montelukast10 11.1 ± 2.9 (N = 9) 10 40 11.4 ± 4.3 (N = 7) 7 Dextromethorphan 5011.1 ± 2.5 (N = 7) 9 100  4.5 ± 2.6 (N = 10) 63

As shown in the above Table 7, it was confirmed that the Sophorae Radixextract of the present invention has an antitussive effect.

EXAMPLE 8 Inhibition of Airway Hyper-Responsiveness

Six week old female BALB/c mice (SLC, Japan) were sensitized byadministering 0.2 mL of a mixture consisting of 10 μg of OVA and alumintraperitoneally at Days 0, 7 and 14, respectively. At 8 and 10 daysafter the sensitization, respectively, 0.7% OVA was sprayed to the micefor 50 min in the form of aerosol by preparing a compressed air underhigh pressure to induce airway hyper-responsiveness. The measurement ofairway hyper-responsiveness was conducted in a barometricplethysmographic chamber (All Medicus, Seoul, Korea) while allowing allthe mice in the chamber free movements and with consciousnesss. The basevalue for airway hyper-responsiveness used was the average value ofmeasurements for 3 min. Then, the mice were inhaled of methacholine for3 min at each concentration of methacholine while gradually increasingits concentration from 2.5 mg/mL to 50 mg/mL and measured the level ofairway hyper-responsiveness at each concentration, respectively.Enhanced pause (Penh) is known as a good index showing the level ofairway resistance and it is calculated based on the following equation1.

Enhanced pause(Penh)=[expiratory time(Te)/relaxation time(RT)−1]×[peakexpiratory flow(PEF)/peak inspiratory flow(PIF)]  [Equation 1]

During the measurement of airway resistance, the average value of Penhis recorded every 10 sec and it is also indicated in Figures every 1min.

As shown in the FIG. 1, it was confirmed that the Sophorae Radix extractof the present invention has an inhibitory activity against airwayhyper-responsiveness.

EXAMPLE 9 Inhibition of Airway Remodeling

Eight to ten week old aseptically treated female BALB/c mice weresensitized by intraperitoneally injecting at Day 1 a mixed solutionconsisting of 500 μg of OVA and 1.0 mg of aluminum hydroxide (firsttreatment), and then inhaled with 2% OVA at Day 11 using an ultrasonicsprayer (second treatment), and finally inhaled with 3% OVA at Days 21,22 and 23 (third treatment). The mice were then allowed to inhale 1% OVAfor the following 6 weeks at intervals of 3 days and constructed anasthma model with induced airway remodeling. In the evening 24 hr priorto the 3rd OVA inhalation, the mice were administered once with theextract (SOS) prepared in Preparation Example 2. The mice wereadministered twice daily with the extract (SOS) prepared in PreparationExample 2 at the 3rd OVA inhalation day (Day 21, 22, and 23), i.e., oneadministration at 1 hr prior to the inhalation and the otheradministration in the evening of the 3rd OVA inhalation day. During the8-week OVA inhalation period for airway remodeling, the mice wereadministered twice daily. The following assays were used for theanalysis of the airway remodeling.

Collagen assay: Method for quantitative analysis of the level of tissuefibrosis

PAS staining: Method for measurement of Goblet cell hyperplasia

Peribronchial trichrome stain: Method of identifying the level offibrosis at the lower bronchus of peribronchial area by staining tissuesusing Masson's trichrome. All bronchi are similar in size based on thelength of the basal membrane and thus 10 bronchi with circular shape areselected and their average value is obtained. The thickness of basalmembrane based on bronchioles having an internal diameter of 150-200 μmare indicated in μm and the ratio is measured by using a computerizedimage analyzer program.

In addition, collected lung tissues are fixed using 10% formaldehydesolution and buried in paraffin blocks. The paraffin blocks were cutinto pieces with a thickness of 1.5 μm to prepare slides. Thus preparedslides were used for observation of histological changes in lungs by H&Estaining.

FIG. 2 shows the effect of the extract prepared in Preparation Example 2(SOS) or montelukast on airway mucus expression in lung tissues ofOVA-sensitized and -challenged mice. Representative PAS-stained sectionsof the lungs. Sampling was performed at 48 hrs after the last challengein saline-inhaled mice with the administration of saline (A),OVA-inhaled mice with the administration of saline (B), OVA-inhaled micewith the administration of montelukast (C), OVA-inhaled mice with theadministration of SOS 50 mg/kg (D), OVA-inhaled mice with theadministration of SOS 100 mg/kg (E), and OVA-inhaled mice with theadministration of SOS 200 mg/kg (F). Bars indicate scale of 50 μm. Barsindicate scale of 50 μm.

FIG. 3 is a histogram showing the effect of the extract prepared inPreparation Example 2 (SOS) or montelukast on airway mucus expression inlung tissues of OVA-sensitized and -challenged mice. The number ofPAS-positive and PAS-negative epithelial cells in individual bronchioleswere counted at 48 hrs after the last challenge in saline-inhaled micewith the administration of saline (SAL+SAL), OVA-inhaled mice with theadministration of saline (OVA+SAL), OVA-inhaled mice with theadministration of montelukast (OVA+MONTE), OVA-inhaled mice with theadministration of SOS 50 mg/kg (OVA+SOS50), OVA-inhaled mice with theadministration of SOS 100 mg/kg (OVA+SOS100), and OVA-inhaled mice withthe administration of SOS 200 mg/kg (OVA+SOS200). Bars represent themeans SEM from 6 independent experiments. #, p<0.05 versus SAL+SAL; *,p<0.05 versus OVA+SAL.

The percentage of airway epithelium staining positively with PAS wasincreased significantly at 48 hrs after OVA inhalation compared to thepercentage after saline inhalation (FIGS. 2 and 3). The increasedpercentage of airway epithelium staining positively with PAS wassignificantly reduced by the administration of SOS or montelukast.

FIG. 4 shows the effect of the extract prepared in Preparation Example 2(SOS) or montelukast on α-smooth muscle expression in lung tissues ofOVA-sensitized and -challenged mice. Representativeimmunohistochemical-stained sections for α-smooth muscle actin of thelungs. Sampling was performed at 48 hrs after the last challenge insaline-inhaled mice with the administration of saline (A), OVA-inhaledmice with the administration of saline (B), OVA-inhaled mice with theadministration of montelukast (C), OVA-inhaled mice with theadministration of SOS 50 mg/kg (D), OVA-inhaled mice with theadministration of SOS 100 mg/kg (E), and OVA-inhaled mice with theadministration of SOS 200 mg/kg (F). Bars indicate scale of 50 μm. Barsindicate scale of 50 μm.

FIG. 5 shows the effect of the extract prepared in Preparation Example 2(SOS) or montelukast on α-smooth muscle expression in lung tissues ofOVA-sensitized and -challenged mice. The area of immunostaining ofα-smooth muscle actin was measured at 48 hrs after the last challenge insaline-inhaled mice with the administration of saline (SAL+SAL),OVA-inhaled mice with the administration of saline (OVA+SAL),OVA-inhaled mice with the administration of montelukast (OVA+MONTE),OVA-inhaled mice with the administration of SOS 50 mg/kg (OVA+SOS50),OVA-inhaled mice with the administration of SOS 100 mg/kg (OVA+SOS100),and OVA-inhaled mice with the administration of SOS 200 mg/kg(OVA+SOS200). Bars represent the means SEM from 6 independentexperiments. #, p<0.05 versus SAL+SAL; *, p<0.05 versus OVA+SAL.

The area of peribronchial α-smooth muscle actin immunostaining wasincreased significantly at 48 hrs after OVA inhalation compared to thearea after saline inhalation (FIGS. 4 and 5). The increased areaof ofperibronchial α-smooth muscle actin immunostaining was significantlyreduced by the administration of SOS or montelukast.

FIG. 6 shows the effect of the extract prepared in Preparation Example 2(SOS) or montelukast on peribronchial fibrosis in lung tissues ofOVA-sensitized and -challenged mice. Representative MassonTrichrome-stained sections of the lungs. Sampling was performed at 48hrs after the last challenge in saline-inhaled mice with theadministration of saline (A), OVA-inhaled mice with the administrationof saline (B), OVA-inhaled mice with the administration of montelukast(C), OVA-inhaled mice with the administration of SOS 50 mg/kg (D),OVA-inhaled mice with the administration of SOS 100 mg/kg (E), andOVA-inhaled mice with the administration of SOS 200 mg/kg (F). Barsindicate scale of 50 μm.

FIG. 7 shows the effect of the extract prepared in Preparation Example 2(SOS) or montelukast on fibrosis in lung tissues of OVA-sensitized and-challenged mice. The area of peribronchial trichrome staining inparaffin-embedded lung was measured at 48 hrs after the last challengein saline-inhaled mice with the administration of saline (SAL+SAL),OVA-inhaled mice with the administration of saline (OVA+SAL),OVA-inhaled mice with the administration of montelukast (OVA+MONTE),OVA-inhaled mice with the administration of SOS 50 mg/kg (OVA+SOS50),OVA-inhaled mice with the administration of SOS 100 mg/kg (OVA+SOS100),and OVA-inhaled mice with the administration of SOS 200 mg/kg(OVA+SOS200). Bars represent mean SEM from 6 independent experiments.Bars represent the means SEM from 6 independent experiments. #, p<0.05versus SAL+SAL; *, p<0.05 versus OVA+SAL.

The area of peribronchial trichrome staining was increased significantlyat 48 hrs after OVA inhalation compared to the area after salineinhalation (FIGS. 6 and 7). The increased area of of peribronchialtrichrome staining was significantly reduced by the administration ofSOS or montelukast.

FIG. 8 shows the effect of the extract prepared in Preparation Example 2(SOS) or montelukast on total lung collagen content of OVA-sensitizedand -challenged mice. The amount of lung collagen was measured using acollagen assay kit. Sampling was performed at 48 hrs after the lastchallenge in saline-inhaled mice with the administration of saline(SAL+SAL), OVA-inhaled mice with the administration of saline (OVA+SAL),OVA-inhaled mice with the administration of montelukast (OVA+MONTE),OVA-inhaled mice with the administration of SOS 50 mg/kg (OVA+SOS50),OVA-inhaled mice with the administration of SOS 100 mg/kg (OVA+SOS100),and OVA-inhaled mice with the administration of SOS 200 mg/kg(OVA+SOS200). Bars represent the means SEM from 6 independentexperiments. #, p<0.05 versus SAL+SAL; *, p<0.05 versus OVA+SAL.

The levels of lung collagen were increased significantly at 48 hrs afterOVA inhalation compared to the area after saline inhalation (FIG. 8).The increased levels of lung collagen were significantly reduced by theadministration of SOS or montelukast.

EXAMPLE 10 Measurement of Cytokines (IL-4, 5 and 13) OVA-specific IgEvia Western Blot Analysis

The concentration of cytokines (IL-4, 5 and 13) in the washed solutionof lung tissues and the concentration of OVA-specific IgE in the serumwere measured by using ELISA kit(cytokine: R&D systems, Abingdon,UK/OVA-specific IgE; BD sciences).

FIG. 9 shows the effect of the extract prepared in Preparation Example 2(SOS) or montelukast on IL-4, IL-5, and IL-13 protein expression in lungtissues of OVA-sensitized and -challenged mice. IL-4, IL-5, and IL-13protein expression were measured at 48 hrs after the last challenge insaline-inhaled mice with the administration of saline (SAL+SAL),OVA-inhaled mice with the administration of saline (OVA+SAL),OVA-inhaled mice with the administration of montelukast (OVA+MONTE),OVA-inhaled mice with the administration of SOS 50 mg/kg (OVA+SOS50),OVA-inhaled mice with the administration of SOS 100 mg/kg (OVA+SOS100),and OVA-inhaled mice with the administration of SOS 200 mg/kg(OVA+SOS200). Results were similar in 8 independent experiments.

Western blot analysis revealed that levels of IL-4, IL-5, and IL-13protein in lung tissues were increased significantly at 48 hrs after OVAinhalation compared to the levels after saline inhalation (FIG. 9). Theincreased levels of these cytokines were significantly reduced by theadministration of SOS or montelukast.

EXAMPLE 11 Toxicity Test on Repeated Oral Administration in Rats

Toxicity tests with repetitive administration were performed using sixweek old specific pathogen free (SPF) SD rats as follows.

Sophorae Radix extract prepared in Preparation Examples 1-3 weresuspended and administered orally in the amount of 2 g/kg for a periodof 2 weeks to rats. Each group contained six animals.

After the administration, the mice were observed with respect to theirdeath, clinical symptoms, change in body weight, hematological andhematobiochemical tests were performed. The mice were then autopsied toexamine any abnormalties in their abdominal and pectoral organs withnaked eyes. The results showed that all the mice survived and there wereno particular signs in terms of clinical symptoms. Further, there wereno significant findings in toxicity test in hematological andhematobiochemical tests as well as in autopsies.

Therefore, it was confirmed that the Sophorae Radix extract of thepresent invention is a safe material which does not exhibit any toxicityin rats up to the dosage of 2,000 mg/kg.

FORMULATION EXAMPLE 1 Preparation of Tablets

Sophorae Radix extract of the present invention with the followingcomposition was formulated in tablets for oral administration by usingwet granulation and dry granulation methods.

[Composition]

Sophorae Radix extract 250 mg, light anhydrous silicone dioxide 10 mg,magnesium stearate 2 mg, microcrystalline cellulose 50 mg, sodium starchglycolate 25 mg, corn starch 113 mg, adequate amount of anhydrousethanol.

FORMULATION EXAMPLE 2 Preparation of Ointments

Sophorae Radix extract of the present invention with the followingcomposition was formulated in ointments.

[Composition]

Sophorae Radix extract 7 g, cetyl palmitate 20 g, cetanol 40 g, stearylalcohol 40 g, isopropyl myristate 80 g, sorbitan monostearate 20 g,polysorbate 60 g, propyl parahydoxybenzoate 1 g, methylparahydoxybenzoate 1 g, adequate amount of phosphate and purified water

FORMULATION EXAMPLE 3 Preparation of Injectables

Sophorae Radix extract of the present invention with the followingcomposition was formulated in injectables.

[Composition]

Sophorae Radix extract 50 mg, mannitol 180 mg, Na₂HPO₄ 25 mg, injectabledistilled water 2,970 mg

FORMULATION EXAMPLE 4 Preparation of Topical Agents

Sophorae Radix extract of the present invention with the followingcompositions was formulated in topical agents.

[Composition 1]

Sophorae Radix extract 0.3 g, sodium polyacrylate 1.3 g, glycerine 3.6g, hydroxy aluminum 0.04 g, methyl parabene 0.2 g, water 14 g.

[Composition 2]

Sophorae Radix extract 0.6 g, propylene glycol 1.6 g, liquid paraffin0.8 g, isopropyl myristate 0.4 g, acrylic adhesive 1430 g, water 16.4 g.

FORMULATION EXAMPLE 5 Preparation of Troches

Sophorae Radix extract of the present invention with the followingcomposition was formulated in troches.

[Composition]

Sophorae Radix extract 1 g, white sugar 50 g, gelatin 3 g, glycerine 10g, acacia gum 1 g, adequate amount of water

FORMULATION EXAMPLE 6 Preparation of Syrups

Sophorae Radix extract of the present invention with the followingcomposition was formulated in syrups.

[Composition]

Sophorae Radix extract 2 g, saccharin 0.8 g, sugar 25.4 g, glycerine 8.0g, flavor 0.04 g, ethanol 4.0 g, sorbic acid 0.4 g, adequate amount ofdistilled water

INDUSTRIAL APPLICABILITY

As stated above, the Sophorae Radix extract of the present invention,unlike the conventional synthesized pharmaceutical drugs which haveshown effective only in limited cases of respiratory diseases model,have exhibited excellent pharmaceutical efficacies in overallrespiratory diseases model. That is, the Sophorae Radix extract of thepresent invention has excellent effect of inhibiting airway contraction,respiratory infections, 5-lipoxygenase activity, phosphodiesterase 4activity, airway hyper-responsiveness and airway remodeling;antagonistic activity against leukotriene D4; and antitussive effect,thus being useful for prevention and treatment of respiratory diseases,such as asthma, acute and chronic bronchitis, allergic rhinitis, acutelower respiratory infections (bronchitis, bronchiolitis, etc.), acuteupper respiratory infections (sphagitis, tonsillitis, laryngitis).

All documents mentioned herein are fully incorporated herein byreference in their entirety.

While the invention has been described with reference to specificembodiments, modifications and variations of the invention may beconstructed with departing from the scope of the invention, which isdefined in the following claims.

1. A pharmaceutical drug for prevention and treatment of respiratorydiseases comprising Sophorae Radix extract as an active ingredient. 2.The pharmaceutical drug according to claim 1, wherein said SophoraeRadix extract is prepared by extracting Sophorae Radix with water or analcohol solution followed by purifying with water-saturated low-gradealcohol or a nonpolar solvent.
 3. The pharmaceutical drug according toclaim 1, wherein said Sophorae Radix extract is prepared by extractingSophorae Radix with water or water-saturated low-grade alcohol or anonpolar solvent followed by purification using the same.
 4. Thepharmaceutical drug according to claim 2, wherein said low-grade alcoholhas carbon atoms of from 1 to 6 and said nonpolar solvent isethylacetate, dichloromethane, chloroform, carbon tetrachloride ormethylethylketone.
 5. The pharmaceutical drug according to claim 1,wherein said respiratory diseases are asthma, acute or chronicbronchitis, allergic rhinitis, acute upper respiratory infection andacute lower respiratory infection.
 6. The pharmaceutical drug accordingto claim 1, wherein said respiratory diseases are caused by airwaycontraction, respiratory infection, 5-lipoxygenase, phosphodiesterase 4,airway hyper-responsiveness or airway remodeling, leukotriene D4, andantitussive effect.
 7. The pharmaceutical drug according to claim 3,wherein said low-grade alcohol has carbon atoms of from 1 to 6 and saidnonpolar solvent is ethylacetate, dichloromethane, chloroform, carbontetrachloride or methylethylketone.